The present invention relates to a heterocycle-containing carboxylic acid derivative and a drug containing said derivative.
Retinoic acid is an essential substance for the growth or life support of mammals including humans. It is known to have various effects, for example, upon ontogenesis, it serves as a morphogenetic factor, while in the adult body, it has effects on the differentiation and proliferation of cells. Described specifically, it is known to take part in the keratinization reaction, hair formation, sebaceous gland function or the like in the epidermis; the metabolism of bone or cartilage in the connective tissue; immunnomodulatory in the immune system; differentiation of neurocytes in the nerve system; differentiation and proliferation of hemocytes in the blood system; and lipid metabolism, mineral metabolism and basal metabolism. A wide variety of physiological effects of retinoic acid as exemplified above are exhibited by its various control mechanisms such as expression regulation of a transcription activator through an intranuclear retinoid receptor (RARs, RXRs) family, regulation of the hormone secretion or function at the target organ, regulation of growth factors such as EGF receptor or TGFA, expression regulation of enzymes such as collagenase, tissue plasminogen activator or tyrosine kinase, and production regulation of cytokine such as IL-6.
In recent years, the relationship between these physiological effects of retinoic acid and various morbid conditions has come to be apparent. Particularly in some cancers typified by acute myelocytic leukemia, differentiation and induction treatment by using all-trans retinoic acid has drawn attentions as a novel therapeutic method of these cancers.
Retinoic acid, however, is found to involve various problems such as appearance of resistance caused by induction of P450 and expression of side effects due to its accumulation. Under such situations, there is accordingly a strong demand for the development of a novel retinoid-related compound, instead of retinoic acid, as a preventive and/or therapeutic for various diseases.
The present invention provides a heterocycle-containing derivative represented by the following formula (I): 
wherein:
A is a group represented by the following formula (i), (ii), (iii) or (iv): 
in which E1, F1, G1, H1, I1, E2, F2, G2, E3, F3 and G3 are the same or different and each independently represents an oxygen atom or a nitrogen, carbon or sulfur atom which may have a substituent; J represents a nitrogen atom which may have a substituent, with the proviso that the cases where in the ring (i) , E1, F1, G1, H1 and I1 each represents a carbon group which may have a substituent, where in the ring (ii), E2, F2 and G2 each represents a carbon-group which may have a substituent, and where in the ring (iii), E3, F3 and G2 each represents a carbon group which may have a substituent are excluded; R1, R2 and R3 are the same or different and each independently represents a hydrogen atom, a halogen atom, a lower alkyl group which may have a substituent, a cycloalkyl group which may have a substituent, a lower alkoxy group which may have a substituent, a cycloalkyloxy group which may have a substituent, an aryl group which may have a substituent, a heteroaryl group which may have a substituent, an aryloxy group which may have a substituent, a heteroaryloxy group which may have a substituent, a cycloalkylalkyl group which may have a substituent, an arylalkyl group which may have a substituent, a heteroarylalkyl group which may have a substituent, a cycloalkylalkyloxy group which may have a substituent, an arylalkyloxy group which may have a substituent, a heteroarylalkyloxy group which may have a substituent, an alkenyl group which may have a substituent or an alkynyl group which may have a substituent, or R1 and R2 are coupled together to form a cycloalkylene group which may have a substituent, a carbon group forming said cycloalkylene group may be substituted by a sulfur atom, an oxygen atom, a sulfinyl group, a sulfonyl group or  greater than NR4, R4 representing a hydrogen atom or a lower alkyl group, and n2 stands for 1 to 3;
B is a heteroarylene group which may have a substituent, an arylene group which may have a substituent, or a group represented by the following formula: 
wherein R5 represents a hydrogen atom or a lower alkyl group; R6 and R7 are the same or different and each independently represents a hydrogen atom, a lower alkyl group or a halogen atom and m1 stands for 1 to 3, m2 stands for 0 to 2; R8 to R15 are the same or different and each independently represents a hydrogen atom or a lower alkyl group; X represents an oxygen atom, a sulfur atom or  greater than NR4, R4 having the same meaning as defined above;
D is an arylene group which may have a substituent, a heteroarylene group which may have a substituent, or a group represented by the formula: xe2x80x94CRxe2x95x90CR7xe2x80x94, R6 and R7 having the same meanings as defined above;
n1 stands for 0 or 1;
M is a hydroxyl group, a lower alkoxy group or a group represented by the following formula: xe2x80x94NR16R17 in which R16 and R17 are the same or different and each independently represents a hydrogen atom, a hydroxyl group, a lower alkyl group, a lower alkoxy group, a hydroxyalkyl group, an aryl group or a heteroaryl group or R16 and R17 are coupled together with the adjacent nitrogen atom to form an oxygen- or sulfur-containing ring; and
 represents a single bond or double bond; or a physiologically acceptable salt thereof.
In the definition of the formula (I), examples of the group represented by A include: 
wherein R1 and R2 have the same meanings as defined above; R18 to R36 are the same or different and each independently represents a hydrogen atom, a lower alkyl group or a phenyl group. Among them, preferred are the groups represented by the following formulae: 
wherein R1 and R2 and R18 to R28 have the same meanings as defined above.
Preferred examples of the group represented by B include heteroarylene groups each of which may have a substituent, groups represented by the formula: xe2x80x94CONHxe2x80x94 or groups represented by the formula: xe2x80x94CR6xe2x95x90CR7xe2x80x94 in which R6 and R7 have the same meanings as defined above.
In the above definition, B embraces the group represented by the following formula: 
D embraces the group represented by the formula xe2x80x94(CR6xe2x95x90CR7)m1xe2x80x94 in which R6, R7 and m1 have the same meanings as defined above; and A embraces the groups represented by the following formulae: 
wherein R1 and R2have the same meanings as defined above, and E, F, G, H and I are the same or different and each independently represents a nitrogen or carbon atom which may have a substituent and at least one of E, F, G, H and I represents a nitrogen atom.
The halogen atom represented by R1, R2, R3, R6 or R7 in the definition of the formula (I) means a fluorine, chlorine, bromine or iodine atom. Examples of the lower alkyl group which is represented by R1 to R3 and may have a substituent include linear or branched C1-6 alkyl groups such as methyl, ethyl, propyl, isopropyl, butyl, isobutyl, sec-butyl, tert-butyl, amyl, isopentyl and neopentyl groups. Examples of the substituent which the lower alkyl group may have include a halogen atom, a lower alkoxy group, a cycloalkyl group, an alkenyl group, an alkynyl group, an aryl group, a heteroaryl group, an arylalkyl group, a heteroarylalkyl group and xe2x80x94NR40R41 in which R40 and R41 are the same or different and each independently represents a hydrogen atom, a lower alkyl group or a lower alkoxy group. Among them, methyl, ethyl, propyl and isopropyl groups are preferred. The lower alkyl group represented by R4 to R36 means any one of the above-exemplified lower alkyl groups.
Examples of the cycloalkyl group which is represented by R1 to R3 and may have a substituent include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl and cycloheptyl groups which may have a substituent. Examples of the substituent which the cycloalkyl group may have include a lower alkyl group, a halogen atom, a lower alkoxy group, a cycloalkyl group, an alkenyl group, an alkynyl group, an aryl group, a heteroaryl group, an arylalkyl group, a heteroarylalkyl group and xe2x80x94NR40R41 in which R40 and R41 have the same meanings as defined above. Among them, a lower alkyl group such as methyl, ethyl and isopropyl, a lower alkoxy group such as methoxy and ethoxy and halogen atoms such as fluorine and chlorine are preferred.
Examples of the lower alkoxy group which is represented by R1 to R3 and may contain a substituent include linear or branched C16 alkoxy groups such as methoxy, ethoxy, n-propoxy, isopropoxy and n-butoxy groups. These lower alkoxy groups may each contain any one of the above-exemplified substituents.
Examples of the cycloalkyloxy group which is represented by R1 to R3 and may contain a substituent include C3-7, cycloalkyloxy groups such as cyclopropyloxy, cyclobutyloxy, cyclopentyloxy, cyclohexyloxy and cycloheptyloxy. These cycloalkyloxy groups may each contain any one of the above-exemplified substituents.
Examples of the aryl group which is represented by R1 to R3 and may contain a substituent include phenyl, 1-naphthyl and 2-naphtyl groups. These aryl groups may each contain any one of the above-exemplified substituents.
Examples of the heteroaryl group which is represented by R1 to R3 and may contain a substituent include groups derived from a heterocyclic ring such as pyridyl, thiazole, oxazole, pyrimidyl, pyrrole, pyrazole, imidazole, furyl and thienyl. These heteroaryl groups may each contain any one of the above-exemplified substituents.
Examples of the aryloxy group which is represented by R1 to R3 and may contain a substituent include phenyloxy and naphthyloxy groups. These aryloxy groups may each contain any one of the above-exemplified substituents.
Examples of the heteroaryloxy group which is represented by R1 to R3 and may contain a substituent include pyridyloxy, thiazolyloxy, oxazolyloxy, pyrimidyloxy, pyrroleoxy, pyrazolyloxy, imidazolyloxy, furyloxy and thienyloxy groups. These aryl groups may each contain any one of the above-exemplified substituents.
Examples of the cycloalkylalkyl group which is represented by R1 to R3 and may contain a substituent include cyloalkylalkyl groups such as cyclopropylmethyl. cyclopropylethyl, cyclobutylmethyl, cyclobutylethyl, cyclopentylmethyl, cyclopentylethyl, cyclohexylmethyl, cyclohexylethyl, cycloheptylmethyl and cycloheptylethyl. These cyloalkylalkyl groups may each contain any one of the above-exemplified substituents.
Examples of the arylalkyl group which is represented by R1 to R3 and may contain a substituent include phenylmethyl, phenylethyl, 1-naphthylmethyl, 1-naphthylethyl, 2-naphthylmethyl and 2-naphthylethyl groups. These arylalkyl groups may each contain any one of the above-exemplified substituents.
Examples of the heteroarylalkyl group which is represented by R1 to R3 and may contain a substituent include groups derived from a heterocyclic ring such as pyridylmethyl, pyridylethyl, thiazolylmethyl, thiazolylethyl, oxazolylmethyl, oxazolylethyl, pyrimidylmethyl, pyrimidylethyl, pyrrolemethyl, pyrroleethyl, pyrazolylmethyl, pyrazolylethyl, imidazolylmethyl, imidazolylethyl, furylmethyl, furylethyl, thienylmethyl and thienylethyl groups. These heteroarylalkyl groups may each contain any one of the above-exemplified substituents.
Examples of the cycloalkylalkyloxy group which is represented by R1 to R3 and may contain a substituent include cyclopropylmethyloxy, cyclopropylethyloxy, cyclobutylmethyloxy, cyclobutylethyloxy, cyclopentylmethyloxy, cyclopentylethyloxy, cyclohexylmethyloxy, cyclohexylethyloxy, cycloheptylmethyloxy and cycloheptylethyloxy groups. These cycloalkylalkyloxy groups may each contain any one of the above-exemplified substituents.
Examples of the arylalkyloxy group which is represented by R1 to R3 and may contain a substituent include phenylmethyloxy, phenylethyloxy, 1-naphthylmethyloxy, 1-naphthylethyloxy, 2-naphtylmethyloxy and 2-naphthylethyloxy groups. These arylalkyloxy groups may each contain any one of the above-exemplified substituents.
Examples of the heteroarylalkyloxy group which is represented by R1 to R3 and may contain a substituent include groups derived from a heterocyclic ring such as pyridylmethyloxy, pyridylethyloxy, thiazolylmethyloxy, thiazolylethyloxy, oxazolylmethyloxy, oxazolylethyloxy, pyrimidylmethyloxy, pyrimidylethyloxy, pyrrolemethyloxy, pyrroleethyloxy, pyraz olylmethyloxy, pyrazolylethyloxy, imidazolylmethyloxy, imidazolylethyloxyv, furylmethyloxyv, furylethyloxy, thienylmethyloxy and thienylethyloxy groups. These heteroarylalkyloxy groups may each contain any one of the above-exemplified substituents.
Examples of the alkenyl group which is represented by R1 to R3 and may contain a substituent include linear or branched C2-6 alkenyl groups, for example, those having a double bond such as ethenyl, propenyl, butenyl, pentenyl and hexenyl. These alkenyl groups may each contain any one of the above-exemplified substituents.
Examples of the alkynyl group which is represented by R1 to R3 and may contain a substituent include linear or branched C2-6 alkynyl groups, for example, those having a triple bond such as ethynyl, propynyl, butynyl, pentynyl and hexynyl. These alkynyl groups may each contain any one of the above-exemplified substituents.
As the cycloalkylene group which is formed by R1 and R2 and may contain a substituent, 5 to 7-membered cycloalkylene groups are preferred. Specific examples include the following groups: 
These cycloalkylene groups may each contain any one of the above-exemplified substituents.
The carbon atom forming the above-exemplified cycloalkylene group may be substituted with a sulfur atom, an oxygen atom, a sulfinyl group, a sulfonyl group or  greater than NR4 in which R4 has the.same meaning as defined above. Specific examples include the following groups: 
wherein R4 has the same meaning as defined above.
The above heterocyclic rings may each contain a substituent, for example, a lower alkyl group such as methyl or ethyl, a halogen atom or a lower alkoxy group.In the definition of B or D, examples of the heteroarylene group which may contain a substituent include divalent groups derived from a heterocyclic ring such as pyridylene, thiazolene, oxazolene, pyrimidylene, pyrrolene, pyrazolene, imidazolene, furylene and thienylene group. These heteroarylene groups may each contain any one of the above-exemplified substituents.
In the definition of B or D, examples of the arylene group which may contain a substituent include phenylene and naphthylene groups. These arylene groups may each contain any one of the above-exemplified substituents.
As the lower alkoxy groups represented by M, R16 or R17, those defined in the above R1 to R3 can be used.
As the hydroxyalkyl groups represented by R16 or R17, the above-exemplified lower alkyl group, any one of carbon atoms of said lower alkyl group containing one to three hydroxyl groups, can be used. As the lower alkyl, aryl and heteroaryl groups, those exemplified above, respectively can be used.
In the definition of R16 and R17, examples of the ring which is formed by R16 and R17 together with the adjacent nitrogen atom and may contain an oxygen atom or sulfur atom include the following rings. 
The term physiologically acceptable salts as used herein means a xe2x80x9cconventionally used nontoxic saltxe2x80x9d. Examples include inorganic acid salts such as hydrochloride, hydrobromide, sulfate and phosphate; organic acid salts such as acetate, maleate, tartrate, methanesulfonate, benzenesulfonate and toluenesulfonate; and salts with an amino acid such as arginine, asparatic acid or glutamic acid. Metal salts of Na, K, Ca and Mg are also embraced by the physiologically acceptable salt of the present invention.
A description will next be made of the typical preparation processes of the invention compound.
Preparation processs 1 
wherein A, D and M have the same meanings as defined above, R50 and R51 individually represent a hydrogen atom or a lower alkyl group, R52 represents a hydrogen atom, a lower alkyl group, an alkenyl group, an alkynyl group, a cycloalkyl group, a cyclokalkylalkyl group, a lower alkoxyalkyl group, an aryl group, a heteroaryl group, an arylalkyl group or a heteroarylalkyl group and Y represents a halogen atom.
A compound of the formula (3) can be obtained by reacting an aldehyde of the formula (1) with an organic metal reagent and then oxidizing the resulting alcohol derivative. As the organic metal reagent for obtaining the alcohol derivative, Grignard reagent, organic lithium reagent or the like can be used. As a solvent, hexane, diethyl ether, tetrahydrofuran or the like can be used. The reaction temperature falls within a range of from xe2x88x9278xc2x0 C. to the boiling point of the solvent, with a range of from xe2x88x9278xc2x0 C. to room temperature being preferred.
As the oxidation reaction, Swern oxidation, manganese dioxide oxidation or chromic acid oxidation can be employed.
A compound represented by the formula (5) can be obtained by reacting the ketone derivative (3) with an aldehyde of the formula (4) in the presence of a catalytic amount of a base and then subjecting the resulting alcohol derivative to dehydration reaction in the presence of an acid. Examples of the base for the preparation of the alcohol derivative include alkali hydroxides such as sodium hydroxide and potassium hydroxide. Examples of the solvent include methanol, ethanol, propanol, tetrahydrofuran and N,N-dimethylformamide. The reaction temperature falls within a range of from 0xc2x0 C. to the boiling point of the solvent, with a range of from 20 to 40xc2x0 C. being preferred.
Example of the acid used for the dehydration reaction include hydrochloric acid, sulfuric acid, p-toluenesulfonic acid, trifluoroacetic acid, oxalic acid and phosphoric acid. Exemplary solvents include ethers such as diethyl ether, tetrahydrofuran, 1,4-dioxane and 1,2-dimethoxyethane and aromatic hydrocarbons such as benzene, toluene and xylene. The reaction is effected at 0xc2x0 C. to the boiling point of the solvent. In some cases, Compound (6) can be obtained directly from Compound (5) without dehydration reaction.
Compound (6) can be introduced into Compound (8) by acting a catalytic amount of a base to Compound (6) with a nitro compound represented by the formula (7) as a solvent (in the case where the compound is sparingly soluble, tetrahydrofuran, methanol, ethanol or the like is added as needed). Exemplary bases include N-benzyltrimethylammonium hydroxide, triethylamine and diisopropylamine. The reaction is effected at a temperature range of from 0xc2x0 C. to the boiling point of the solvent, with a temperature range of from 0C to room temperature being preferred.
Compound (9) can be obtained by subjecting Compound (8) successively to Nef reaction and ketalization. Ketalization is attained by the addition of a mineral acid such as sulfuric acid or hydrochloric acid in methanol. The reaction temperature falls within a range of from xe2x88x9278xc2x0 C. to the boiling point of the solvent, with a range of from xe2x88x9240xc2x0 C. to room temperature being preferred.
A pyrrole derivative (10) can be obtained by acting a primary amine (14) to Compound (9). Any solvent can be used insofar as it is inert to the reaction. Preferred are aromatic hydrocarbon solvents such as benzene, toluene and xylene, ether solvents such as tetrahydrofuran and 1,2-dimethoxyethane and alcohol solvents such as methanol and ethanol. The reaction is allowed to proceed in the presence of an acid in the above-exemplified solvent. As the acid, that which also serves as a dehydrating agent, for example, hydrochloric acid, sulfuric acid, glacial acetic acid or polyphosphoric acid can be used. The reaction can also be effected by using an acid such as glacial acetic acid as a solvent.
When Compound (9) is reacted using an ammonium salt such as ammonium acetate, ammonium hydrochloride or ammonium sulfate in the presence of an acid, a pyrrole derivative (11) can be obtained. Compound (10) can be obtained by acting a halide (15) to Compound (11) in the presence of a base. Exemplary bases include alkali metal compound such as potassium carbonate, sodium hydride and potassium hydride and metal alkoxides such as sodium methoxide, sodium ethoxide and potassium-tert-butoxide. Examples of the solvent include N,N-dimethylformamide, tetrahydrofuran and 1,2-dimethoxyethane. The reaction temperature ranges from 0xc2x0 C. to the boiling point of the solvent.
Compound (9) can be introduced into its furan derivative (12) by acting an acid on it. Illustrative acids include sulfuric acid and polyphosphoric acid. The reaction is effected at 0xc2x0 C. to 100xc2x0 C. Compound (9) can be introduced into a thiophene derivative (13) by acting, on Compound (9), a sulfide such as phosphorus pentasulfide or hydrogen sulfide. As the solvent, an aromatic hydrocarbon such as benzene, toluene or xylene or pyridine is used and the reaction is effected at a temperature range of from 0xc2x0 C. to the boiling point of the solvent, preferably from 50xc2x0 C. to the boiling point of the solvent.
Preparation Process 2 
wherein A, D, M, Y, R50, R51 and R52 have the same meanings as defined above.
A compound represented by the formula (16) can be prepared by reacting an aldehyde of the formula (1) with an organic metal reagent as in Preparation Example 1 and then oxidizing the resulting alcohol derivative.
Compound (17) can be obtained by reacting Compound (16) with an aldehyde of the formula (4) in the presence of a base through the use of 3-benzyl-5-(2-hydroxyethyl)-4-methylthiazolium chloride. Examples of the base include potassium carbonate, pyridine and triethylamine, while examples of the solvent include alcohol solvents such as methanol and ethanol, and N,N-dimethylformamide. The reaction temperature ranges from room temperature to the boiling point of the solvent, with a range of from 50xc2x0 C. to the boiling point of the solvent being preferred.
Alternatively, Compound (17) can be obtained in a similar manner by reacting the aldehyde of the formula (1) with a compound of the formula (16xe2x80x2).
From a xcex3-diketone of the formula (17), its pyrrole derivative (17), furan derivative (20) or thiophene derivative (21) can be obtained in a similar manner to the conversion of Compound (9) to Compound (10), (12) or (13), respectively, in Preparation Process 1.
Preparation Process 3 
wherein A, D, M, Y and R50 have the same meanings as defined above and L represents S or  greater than NR53 in which R53 represents a hydrogen atom or a lower alkyl group.
A ketone derivative (3) is halogenated at its a position and then the resulting halogenated product (22) is reacted with a thioamide or amidine represented by the formula (23), whereby the corresponding thiazole or imidazole derivative represented by the formula (24) can be prepared. Exemplary halogenating reagents include bromine, copper bromide, N-bromosuccimide, chlorine, N-chlorosuccimide and iodine. The conversion into a heterocyclic compound is attained at the reaction temperature ranging from 0xc2x0 C. to the boiling point of the solvent in the presence of a base such as pyridine, triethylamine or potassium carbonate in a solvent such as methanol, ethanol, isopropanol, tetrahydrofuran or N,N-dimethylformamide.
Preparation Process 4 
wherein A, D, M, Y, R50 and R52 have the same meanings as defined above.
A diketone derivative (26) can be obtained by reacting a ketone derivative (3) with an acid halide (25) in the presence of a base. As the base, lithium diisopropylamide or lithium bistrimethyl silylamide brings about good results. Examples of the solvent usable in the above reaction include ether solvents such as diethyl ether, tetrahydrofuran and 1,2-dimethoxyethane. The reaction temperature ranges from xe2x88x9278xc2x0 C. to the boiling point of the solvent, with xe2x88x9278xc2x0 C. to room temperature being preferred.
The diketone derivative (26) is reacted with hydrazine hydrate, whereby a pyrazole derivative (27) can be obtained. This reaction can be accelerated by the addition of hydrochloric acid, sulfuric acid, acetic acid, polyphosphoric acid or the like as a dehydrating agent. Any reaction solvent can be used insofar as it is principally inert to hydrazine. Examples include alcohol solvents such as methanol, ethanol and isopropanol, aromatic hydrocarbon solvents such as benzene, toluene and xylene, N,N-dimethylformamide and dimethylsulfoxide. Acids such as acetic acid can also be used as a solvent. The reaction temperature ranges from 0xc2x0 C. to the boiling point of the solvent, with a range of from room temperature to the boiling point of the solvent being preferred. A pyrazole compound (29) can be obtained by carrying out the same reaction by using a hydrazine represented by the formula (28) or by reacting the pyrazole derivative (27) with a halide (15) in a similar manner to preparation process 1 and separating the resulting isomer by crystallization or column chromatography.
Preparation Process 5 
wherein A, D and M have the same meanings as defined above and R54 represents a lower alkyl group.
A compound represented by the formula (31) can be obtained by alkali hydrolysis of an ester derivative represented by the formula (30). This reaction is carried out in a mixture of water and an alcohol such as methanol, ethanol or propanol, tetrahydrofuran or 1,4-dioxane in the presence of an alkali hydroxide such as sodium hydroxide or potassium hydroxide used in an excess amount. The reaction temperature falls within a range of room temperature to the boiling point of the solvent mixture, preferably within a range of from room temperature to 60xc2x0 C. An amide compound represented by the formula (33) can be obtained by converting a carboxylic acid as compound (31) into corresponding acid halide, acid azide or acid anhydride in a manner known to date, followed by the reaction with an amine compound represented by the formula (32).
Preparation Process 6 
wherein A, D and M have the same meanings as defined above.
A compound of the formula (36) can be obtained by converting a carboxylic acid of the formula (35) into the corresponding acid halide, acid azide or acid anhydride in a manner known to date and then reacting an amine compound of the formula (34) therewith.
Furthermore, other compounds of the formula (1) according to the present invention can also be obtained by converting, in a manner known to date, the group of the compound prepared by any one of the processes described as Preparation Processes 1 to 6 or another process, said group being represented by the formula:
xe2x80x94Dxe2x80x94COxe2x80x94M
wherein D and M have the same meanings as defined above.
In the case where the group represented by the formula: xe2x80x94Dxe2x80x94COxe2x80x94M is derived from a carboalkoxy-containing benzoate compound, it can be converted into its free carboxylic acid or physiologically acceptable salt thereof by alkali hydrolysis. The alkali hydrolysis is effected in a mixture of water with an alcohol such as methanol, ethanol or propanol, tetrahydrofuran or 1,4-dioxane in the presence of an alkali hydroxide such as sodium hydroxide or potassium hydroxide used in an excess amount at a temperature ranging from room temperature to the boiling point of the solvent.
The effects of the invention compound will next be described by Pharmacological Experimental Examples.
Receptor Binding Assay by Using Human Promyelocytic Leukemia Cells HL60
It is known that there exists a receptor for all-trans retinoic acid (Retinoic acid receptor: RAR) in the nucleus of HL60 cells [Clara Nervi et al., Proc. Natl. Acad. Sci., 86 5854(1989)]. Specific binding of all-trans retinoic acid with PAR was found using a fraction extracted from the nucleus of HL60 and the binding capacity of each compound with PAR was studied by measuring the binding inhibition rate.
The nucleus-extracted fraction was prepared as described below.
In 15 ml of Solution A [5 mM sodium phosphate (pH 7.4), 10 mM monothioglycerol, 10% v/v glycerol, 1 mM phenylmethyl sulfonyl fluoride (PMSF), 10 xcexcg/ml of aprotinin and 25 xcexcg/ml of leupeptin], HL60 cells (5xc3x97108) were suspended. The resulting suspension was homogenized, followed by centrifugal separation to remove the supernatant. The precipitate so obtained was suspended in 15 ml of Solution B [10 mM tris-HCl (Tris-HCl) (pH 8.5), 10 mM monothioglycerol, 10% (v/v) glycerol, 1 mM PMSF, 10 xcexcg/ml aprotinin, 25 xcexcg/ml leupeptin and 0.4 M KCl]. After being allowed to stand at 4xc2x0 C. for one hour, the resulting suspension was subjected to ultracentrifugation at 100,000xc3x97g and 4xc2x0 C. for one hour. The supernatant so obtained was refrigerated at xe2x88x9280xc2x0 C. as a nucleus-extracted fraction until practical use (METHODS IN ENZYMOLOGY, 189, 248).
Receptor binding assay was carried out as follows:
To a polypropylene-made 96-well plate, 180 xcexcl of the extracted fraction and 10 xcexcl of all-trans retinoic acid or a diluted invention compound were added, followed by the addition of 10 xcexcl of 10 nM 3H-all-trans retinoic acid. The resulting mixture was allowed to stand at 4xc2x0 C. for 16 hours. To the reaction mixture was added a 3% charcoal-0.3% dextran solution and the resulting mixture was centrifuged to separate free 3H-all-trans retinoic acid. The count of the supernatant was determined by a scintillation counter. The specific binding with PAR was determined by deducting the above count from a count at the time when all-trans retinoic acid was added in an amount excessive by 200 times as a value of nonspecific binding. The compounds which will be described below suppressed the binding of 3H-all-trans retinoic acid according to the concentration. In addition, the 50% inhibition concentration of each compound was calculated and is shown in Table 1.
Differentiation and Induction Effects on HL60 Cells
It is known that human-derived promyelocytic leukemia cell strains HL60 differentiate into the corresponding granulocytic cells in the presence of all-trans retinoic acid [Breitman, T. Selonick., S. and Colling, S. Proc. Natl. Acad. Sci., 77, 2936(1980)]. In general, cells after differentiation have come to express specific differentiated antigens on their surface. When the HL60 cells are differentiated into granulocytic cells, CD11b which is a differential antigen between granulocytes and monocytes is expressed on the surface of the cells [Fontana, J A., Reppuci, A. Durham, J P. and Mirand, D. Cancer Res. 46, 2469-2473(1986)]. Effects of the invention compound on the differentiation and induction into granulocyte cells were studied by making use of the above-described phenomenon.
HL60 cells were cultured and kept in RPMI 1640 (a medium formulated by Rosewell Park Memorial Institute) to which 10% bovine fetal serum, 1 mM sodium pyridinate, 50 xcexcM xcex2-mercaptoethanol, 100 IU/ml of penicillin and 100 pg/ml of streptomycin had been added.
On each well of a 24-well plate, a 1 ml portion of the suspension containing HL60 in an amount of 1xc3x97105 cfu/ml was seeded. The invention compound was then added thereto at varied concentrations, followed by cultivation for 5 days in a 5% CO2-air incubator. After cultivation, the cells of each well were collected in a test tube, to which an FITC-labeled monoclonal antibody against Cd11b, that is, a granulocyte-monocyte specific antigen was added. The cells were then immobilized with 0.2% paraformaldehyde. The existing ratio of CD11b positive cells in the vial HL60 cell group present in each well was determined using flow cytometry [Miller, L J., Schwarting, R. and Springer, T A. J. Immunol. 137, 2891-2900(1986)]. The existing ratios of CD11b positive cells of compounds which will be described below were increased according to the concentration. The concentration at which the existing ratio of the positive cells becomes 33% is defined as ED⅓. The ED⅓ of each compound is calculated and is shown in Table 1.
As is apparent from the above results, compounds according to the present invention have retinoid receptor agonist effects and are therefore expected as medicaments effective against various diseases, which will be described below, as a preventive and/or therapeutic for autoimmune diseases, immunosuppression upon organ transplantation and malignant neoplasm. The use of the compounds of the present invention are not limited to the following diseases.
Various keratosis, psoriasis, acne, leukoplakia, xerodetma pigmentosum.
Various alopecia such as alopecia areata, alopecia seborrheica, melanoderma cachecticorum.
Postmenopausal osteoporosis, senile osteoporosis, cataplectic osteoporosis, diabetic osteopenia, chronic rheumatism osteopenia, renal osteomalacia, ectopic ossification.
Osteoarthritis, glenoidal periarthritis
Autoimmune diseases such as chronic rheumatoid arthritis, multiple sclerosis (MS), systemic lupus erythematosus (SLE), Behcet disease, mycosis fungoides (MF), progressive systemic sclerosis, dermatomyositis (DM) and nodular arteriosclerosis.
Rejection symptoms upon organ transportation
Atopic dermatitis
Asthma (immediate allergy reaction)
Immune function activation in immunodeficiency, cytomegalovirus infectious diseases of a fetus or upon immune depression, opportunistic infection.
Hyperthyroidism
Cancers such as squamous cell carcinoma, cystocarcinoma, pulmonary carcinoma, esophageal carcinoma, cervical carcinoma, large bower/rectum cancer, prostatic cancer, uterocervical carcinoma, mammary carcinoma, neurocytoma, acute promyelocytic leukemia, acute myelocytic leukemia, osteomyelodysplasia, chronic myelocytic leukemia and cutaneous T-cell lymphoma.
Hyperkalemia
Pulmonary fibrosis, hepatic fibrosis, hepatic cirrhosis.
For the administration of the invention compound as A preventive and/or therapeutic for the above-exemplified disease, it may be orally administered as tablets, powders, granules, capsules or syrups; or it may be parenterally administered as a suppository, injection, external preparation or drops.
These preparations for oral or parenteral administration are formulated in a manner known per se in the art by using an ordinarily used pharmaceutically acceptable carrier.
Subcutaneous, intramuscular and intravenous injections and dropping injection are formulated in a manner known per se in the art by adding, if necessary, a pH regulator, buffer, stabilizer and/or solubilizing agent to a main agent, followed by lyophilization as needed.